Sexual transmission is the dominant mode for HIV spread worldwide. Our goal is to develop a subunit vaccine based on the HIV gp120 envelope glycoprotein, which can be delivered via atraumatic mucosal inoculation to elicit durable protective immunity. In order to immunize at mucosal sites, we need an efficient method for delivering gp120 and adjuvant to mucosal immune cells. We constructed fusion proteins consisting of gp120 linked to the Fc portion of IgG2 antibodies. The fusion protein Fc segment binds Fc receptor neonatal (FcRn) on mucosal epithelium, and crosses the epithelial barrier by non-degradative transcytosis to access underlying antigen presenting cells. Adjuvant, in our case muramyl dipeptide (MDP), is conjugated directly to the gp120-Fc fusion protein through modification of terminal sialic acid residues (Env-Fc-MDP). Thus, conjugated adjuvant and antigen are co-delivered to presenting cells. MDP was selected because it increases co-stimulatory molecule and IL-12 expression by dendritic cells, and should enhance T and B cell responses to HIV Env. The potency and durability of gp120 immune responses is increased with a prime/boost strategy where mucosal priming with fusion protein is followed by intramuscular boosting with gp120 plus soluble MDP. Our studies utilize in vitro or mouse models to optimize immunization strategies with fusion protein and adjuvant. Nonhuman primate studies optimize dose and route for immunization using an unrelated fusion protein (HSV-2 glycoprotein D-Fc), then test whether mucosal Env-Fc-MDP followed by intramuscular Env + soluble MDP elicits protection against repetitive, low-dose intrarectal inoculation with SHIV162p3. This project is an interdisciplinary collaboration between Dr. Zhu, expert in mouse models and Fc receptor biology, Dr. Wang, who is a glycobiologist and organic chemist expert in protein glycosylation, and Dr. Pauza who has extensive experience in nonhuman primate models, mucosal immunology and mucosal virus infections. The innovative route for mucosal antigen delivery, use of chemically-conjugated adjuvant to improve T cell responses, and our prime/boost strategy is a safe and potentially effective method for eliciting durable protective immunity against sexual transmission of HIV-1.